Programmed cell death (PCD or apoptosis) is an evolutionarily conserved mechanism essential for the development and maintenance of tissue and organismal homeostasis in multicellular organisms. All available evidence is consistent with the idea that one of the most biologically relevant classes of proteins regulating PCD processes at the effector phase common to all PCD processes is a family of proteins related to the bcl-2 gene product. The Principal Investigator intends to test current models of the action of this family of proteins which invoke regulation of the mitochondria permeability transition (PT) by bcl-2 family members, leading to modulation of early mitochondria events which characterize PCD pathways. The PT is inhibited by nanomolar concentrations of the immunosuppressant peptide cyclosporin A (CsA), as is PCD in a number of assay systems. A large body of data indicates that the only known target of CsA action in mitochondria is CyP-D, a mitochondria matrix cyclophilin whose enzymatic activity is inhibited by CsA and a variety of non-immunosuppressive analogs. The specific focus of this proposal is then to unambiguously resolve basic questions related to: 1) the mechanism of PT inhibition by CsA, 2) the influence of CyP-D on the PT and 3) the role of the PT in PCD and other pathological processes. Three aims are proposed. Aim 1 is to generate CyP-D knockout mice. The second aim is to characterize their phenotypes. Finally, the Principal Investigator will characterize the mice at the cellular level to determine the effects on mitochondrial function.